Comparison of the neuroprotective effects of adrenoceptor drugs in retinal cell culture and intact retina

PURPOSE. The efficacy of β₁,-adrenoceptor (AR)-selective (betaxolol and metoprolol) and nonselective (timolol) antagonists and the α₂-AR agonist UK14,304 as retinal neuroprotectants was compared and contrasted in an in vitro glutamate excitotoxicity model. The ability of UK14,304, brimonidine, and betaxolol to alter glutamate-receptor-induced changes in intracellular calcium ([Ca²⁺]_i) was also determined in isolated retinal neurons and retinal ganglion cells (RGCs) in an intact retina preparation. METHODS. Neuronal survival was measured in mixed retinal cell cultures treated for 24 hours with media containing 100 μM glutamate, with or without the addition of each of the drugs (1-1000 μM). Effects of glutamate on glia were also investigated in a C6 glioma cell line. Glutamate-induced changes in [Ca²⁺]_i with and without UK14,304, and its analogue brimonidine were assessed by calcium-imaging techniques in retinal neurons in culture. The effect of betaxolol on [Ca²⁺]_i was investigated in RGCs in intact rabbit retina. RESULTS. In cell cultures, 10 - 1000 μM glutamate resulted in a dose-dependent loss of neurons, but not of glia. The absence of glutamate toxicity in glia was confirmed in C6 glioma cells. Betaxolol, but not timolol or metoprolol, significantly increased survival (from 52% of control in glutamate-only to 78% with 10 μM betaxolol) after excitotoxic insult. UK14,304 also increased survival (from 62% of control in glutamate only to 109% and 101% of control with 10 and 100 μM UK14,304, respectively). This effect was blocked by the specific α₂-antagonist, yohimbine. Both UK14,304 and brimonidine (10-100 μM) reduced glutamate-induced [Ca²⁺]_i increases in retinal neurons in culture. The actions of the α₂-agonists in reducing glutamate-induced [Ca²⁺]_i increases were reduced by yohimbine (1 μM). Betaxolol (100 μM) reduced N-methyl-o D-aspartate (NMDA)-induced increases of [Ca²⁺]_i in RGCs in intact retina. CONCLUSIONS. Betaxolol reduced glutamate excitotoxicity in retinal neurons in vitro through a mechanism independent of β-AR interactions. UK14,304, acting through α₂-ARs, was also neuroprotective in vitro. The neuroprotective actions of betaxolol and the α₂-agonists on retinal neurons may be due, at least in part, to a direct reduction of glutamate receptor-mediated increases of [Ca²⁺]_i.